The long-term objective of this proposal is to understand how neuropeptides interact in the brain to regulate energy homeostasis. Both leptin and insulin signal the level of energy stores to the brain and activate neuropeptide centers in the hypothalamus which modulate appetite and body weight. This proposal will focus on the melanocortin neuropeptide system which is an important target for leptin in the hypothalamus. Studies will center on the regulation of proopiomelanocortin (POMC) and the POMC-denved peptides, alpha-MSH, gamma-MSH and beta-EP, together with the newly discovered agouti related protein (AGRP) which is synthesized in the hypothalamus and is a potent antagonist of the MSH peptides. a-MSH inhibits feeding and AGRP is an orexigemc peptide which antagonizes the actions of alpha-MSH at specific melanocortin receptors. Although lepthi has been shown to stimulate POMC gene expression in the hypothalamus, nothing is known about the effects of leptin on POMC peptide processing and release. This is an important consideration because POMC is processed to a number of peptides with different biological activities. Little is known about the regulation of AGRP in the hypothalamus which is coexpressed in the same neurons with the orexigenic peptide NPY. Several Aims will focus on the regulation of POMC, AGRP and NPY in the rat by both leptin and insulin and on interactions between the POMC and AGRP neurons themselves which both express melanocortin receptors. Neuropeptide gene expression will be studied by sensitive solution hybridization assays; peptide levels and processing will be studied by gel filtration, HPLC and well characterized RIAs; peptide release will be studied in a well validated in vitro perifusion system. The third Aim will focus on the role of leptin and the POMC peptides in the regulation of the hypothalamic-pituitary-adrenal (HPA) axis, which plays a key role in energy homeostasis, and is intricately related to the obesity syndromes in leptin deficient animals. A major hypothesis to be tested is that alpha-MSH and beta-EP inhibit the HPA response and mediate inhibitory effects of leptin on the HPA axis. In the final Aim, the effects of alpha-MSH overexpression in transgenic mice will be studied with respect to energy homeostasis, neuropeptide gene expression and HPA responses. There is growing evidence that POMC plays a role in human energy homeostasis. Obesity has recently been reported in patients with P0MG and melanocortin receptor mutations. Our proposed studies of P0MG regulation and processing in the rodent should thus have implications for the regulation of food intake and body weight in the human.